Prostaglandins (PG) and leukotrienes (LT) play a critical role in pathophysiology. In particular, inflammation is both initiated and maintained by the overproduction of prostaglandins and leukotrienes in injured cells. Prostaglandins and leukotrienes are biosynthesized on demand from arachidonic acid, a 20-carbon fatty acid that is derived from the breakdown of cell-membrane phospholipids. Prostaglandins and leukotrienes are produced via separate enzyme pathways known as the cyclooxygenase (COX) and lipoxygenase (LO) pathways, respectively.
The first step in the synthesis of prostaglandins is the cyclooxygenase reaction. An enzyme, cyclooxygenase (COX) (also known as prostaglandin H synthase (PGHS)) catalyzes the conversion of arachidonic acid into the endoperoxide PGG2, and endoperoxide PGG2 to PGH2. PGH2 is in turn metabolized by one or more prostaglandin synthase (PGE2 synthase, PGD2 synthase, etc.) to generate the final “2-series” prostaglandins, such as PGE2, PGD2, PGF2a, PGI2, and others, as well as thromboxanes and prostacyclins.
As disclosed in U.S. Pat. No. 6,048,850 (to Young et al.), there are two forms of COX. Cyclooxygenase-1 (COX-1) is “constitutively” expressed in most tissues. It is described as a “housekeeping” enzyme, regulating normal cellular processes, such as gastric cytoprotection, vascular homeostasis, platelet aggregation, and kidney function. Although generally described as “constitutive,” COX-1 also can be induced. Ferraz et al., Gastroenterology, 113(1):195–204 (1997). Further, an increase in COX-1 mediated prostaglandin synthesis, such as PGI2, has been reported as important in preventing gastric lesion formation. Harada et al., J. Lab. Clin. Med., 129(6):620–6 (1997). This may be because prostaglandins help maintain an intact gastric mucosal barrier by increasing secretion of mucus and bicarbonate, maintaining mucosal blood flow, and decreasing acid secretion. Fuller and McKenzie, U.S. Pharmacist, 17:35–36, 41–42, 47–48, 53–55, 87 (1992).
Cyclooxygenase-2 (COX-2) is usually undetectable in most tissues; however, its expression is increased during states of inflammation or, experimentally, in response to mitogenic stimuli. COX-2 is accordingly referred to as “inducible.” It is this inducible COX-2 form that is responsible for prostaglandin overproduction through the COX pathway in response to tissue injury, and stimulation by growth factors and proinflamatory cytokines.
As the COX pathway is the rate-limiting step for prostaglandin synthesis, the COX reaction is the principal target for anti-inflammatory drug action. And it is inhibition of COX activity that accounts for the activity of the non-steroidal anti-inflammatory drugs (NSAIDs), such as aspirin, acetaminophen, ibuprofen, naproxen, indomethacin. These drugs however, are nonselective in their inhibition of COX. Thus, they inhibit the activity of COX-2 in inflammation, which produces a desirable therapeutic effect. But they also inhibit the activity of COX-1 in non-inflamed cells, which interferes with the normal production of prostaglandins necessary for normal physiological functions. COX-1 activity generates protective gastric mucosal prostaglandins, as disclosed in U.S. Pat. No. 6,344,323 (Siefert). Inhibiting such prostaglandin production by COX-1 can produce undesirable side effects, such as renal failure, and gastrointestinal mucosal disorders, for example, gastritis, gastrointestinal bleeding, and ulcers. An estimated 16,500 deaths each year result from gastrointestinal bleeding associated with traditional NSAIDs, as reported by Moskowitz, Consultant, 40:1370 (2000).
In an effort to utilize the benefits of COX-2 inhibition without the side effects of COX-1 inhibition, pharmaceutical compounds have been developed to selectively inhibit the activity of COX-2. For example, celecoxib (marketed by Pharmacia/Pfizer as CELEBREX) and rofecoxib (marketed by Merck as VIOXX) are both selective COX-2 inhibitors. Nimesulide is a Cox-2 inhibitor marketed under the trade name MESULID in Europe. COX-2 selectivity can be quantified by calculating the COX-2/COX-1 IC50 (inhibitor concentration at which 50% inhibition occurs) ratio. Compounds with a ratio less than one can be considered relatively COX-2 selective—the lower the ratio, the higher the COX-2 selectivity.
The literature is replete with reports that selective COX-2 inhibiting compounds are usefull in treating a variety of conditions, which are mediated at least in part by inflammation. For example, COX-2 inhibitors have been reported as useful to treat conditions such as general pain, osteoarthritis and rheumatoid arthritis, Whelton et al., Am J. Ther, 7(3): 159–75 (2000), menstrual pain associated with primary dysmenorrhea, Daniels et al., Obstet Gynecol, 100(2):350–8 (2002), cancers, such as colon cancer, Nagatsuka, et al., Int'l. J. Cancer, 100(5):515–9 (2002), oral cancer, Wang et al., Laryngoscope, 112(5):839–43 (2002), and skin cancer, Lee et al., Anticancer Res., 22(4):2089–96 (2002); Fischer, J. Environ. Pathol. Toxicol. Oncol. 21(2):183–91 ((2002), and Alzheimer's disease, Aisen, J. Pain Symptom Manage., 23(4 Suppl):S35–40 (2002).
The herb Carthamus tinctorious (safflower) is a member of the family Compositae and the sub-family Asteridae, which includes the sunflower, artichoke, tarragon, southernwood, and many ornamentals. Carthamus tinctorious flower petals have traditionally been used in Chinese medicine to improve blood circulation and to treat pain or inflammation. Kee Chang Huang, PHARMACOLOGY OF CHINESE HERBS, 2nd Ed., p.318–20 (1999). Extracts of Carthamus tinctorious flowers have been made, for example a 60% ethanol/40% water extract, and investigated to determine chemical constituents of the plant, such as flavanoids, as well as to determine blood circulation effects caused by the extracts. Li & Che, Yao Xue Xue Bao, 33:626–8 (1998). A composition comprising Echinacea purpourea and one or more of several other plants including Carthamus tinctorious has been disclosed as an inhibitor of PGE2 production in Japanese Patent Application No. 10184282 by Yumiko et al. However, no references are known that disclose that any extract of Carthamus tinctorious , or any other extract from a plant in the Asteridae sub-family, selectively inhibits COX-2 activity. Also, although pharmaceutical compositions of extracts of the seeds of Carthamus tinctorious have been disclosed for the treatment of inflammation, U.S. Pat. No. 6,245,340 (to Youssefyeh), no pharmaceutical compositions including extracts from the flower or other parts of Carthamus tinctorious other than the seeds are known.
In the lipoxygenase pathway, lipoxygenase (such as 5-LO) converts arachidonic acid into a hydroperoxy eicosatetraenoic acid (such as 5-hydroperoxy-eicosatetraenoic acid (5-HPETE)), which is converted to the leukotriene LTA4 that can in turn be converted to leukotrienes LTB4, LTC4, or LTD4. Leukotrienes are implicated in many inflammatory diseases and related conditions including cancer proliferation and also can contribute to the development of gastrointestinal ulcers by contributing to the inflammatory process. Leval et al., Curr. Med. Chem., 9:941–62 (2002); Fiotucci et al., Biochem Pharmacol., 62:1433–8 (2001).
Because of the inflammatory action of leukotriene production, inhibitors of lipoxygenase have been used to inhibit the production of leukotrienes in order to inhibit inflammation. Muller-Peddinghaus, Physiol Pharmacol., 48:529–36 (1997).
The plant Bosswellia serrata (specifically the gum resin of the plant) has been used for the treatment of rheumatoid arthritis and gout by the practitioners of Ayurvedic medicines in the Indian system of medicine, as discussed in EP-A 7559000040. In particular, various extracts of the gum resin containing acids known as boswellic acids have shown anti-inflammatory and anti-arthritic activity in laboratory animals, as well as during clinical trials. Atal et al., Ind. J. Pharm, 12, 59 (1980); Pachnanda et al., Ind. J. Pharm, 13, 63 (1981). Singh et al., established that an alcohol extract of Boswellia serrata gum resin displayed anti-inflammatory activity in carrageenan induced edema in rats and mice and dextran induced edema in rats. Singh et al., Agent and Action, 18:407 (1986). Safayhi et al. showed that an ethanolic extract of Bosswellia serrata inhibited 5-LO product formation. Safayhi et al, Planta Med., 66:110–3 (2000).
The effect of Bosswellia serrata extract on COX-2 activity has been studied, but boswellic acids isolated from the gum resin of Bosswellia serrata were found not to inhibit COX-2 activity. Ammon et al., J. Ethnopharmacol. 38:113–9 (1993); Safayhi et al., J. Pharmacol. Exp. Ther., 261:1143–6 (1992).